Abundant expression of enzymes of fatty acid synthesis occurs for each cycle of lactation and in a subset of aggressive breast cancers. S14 is a hormonally-inducible nuclear protein expressed only in tissues actively synthesizing lipids for use as a fuel (lactating mammary, liver, adipose), and is required for induction of genes coding lipogenic enzymes in response to hormonal and dietary signals. S14 is expressed in human mammary epithelium only during lactation, and we find it is critical for lipid metabolism, growth, and survival of mammary epithelial and lipogenic breast cancer cells. Genes coding S14 and Cyclin D1, a mammary oncogene that is also key for normal mammary development, both reside on the l lq13 cancer amplicon. In addition to S14 gene amplification, we find S14 overexpression in the majority of tumors, as is the case for cyclin D1. S14 expression correlates with that of a key lipogenic enzyme (acetyl-CoA carboxylase) in tumors, and is induced by progestin in breast cancer cells in concert with fatty acid synthase (FAS). This proposal is focused on the hypothesis that increased fatty acid synthesis in both lactating and malignant mammary cells is mediated by S14 and that this is critical for cell proliferation. We propose four specific aims: First, to define the impact of S14 expression on growth and metabolism of cells derived from normal mammary epithelium and from breast cancer. S14 expression and antisense genes will be introduced into HC11 lactating mouse mammary epithelial- and human breast cancer cells. Lipid synthesis, expression of lipogenic enzymes and their mRNAs, and growth/cell cycle effects will be analyzed in tissue culture and in xenografts in nude mice. Second, to define the functional interaction of S14 arid a key transcription factor (SREBP1c) that induces S14 and other genes for lipid synthesis and is also required for survival of lipogenic breast cancer cells. Regulation of FAS gene transcription by S14 and SREBP1c will be assessed by deletion analysis, and protein-DNA interactions will be identified. Third, to determine whether S14 drives mammary lipogenesis in vivo, and whether it acts as an oncogene, either alone or in cooperation with cyclin Dl. MMTV-S14 transgenic mice will be assessed metabolically and for tumorigenesis. In addition, MMTV-S14 mice will be crossed with a MMTV-cyclin D1 strain to determine if metabolism and/or virulence of cyclin D 1-induced tumors are influenced by S14. Fourth, to assess the role of S14 in mammary metabolism and development in mice with mammary-specific S14 gene disruption. These studies will provide a comprehensive view of the role of this novel nuclear protein in both normal mammary physiology and in breast cancer, and are likely to reveal a new class of therapeutic targets.